GABAAReceptor Populations Bind Agonists and Antagonists Differentially and with Opposite Affinities

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Abstract
Pretreatment of synaptosomal membranes with a diazo-coupling reagent and the presence of Cl- ions were used to differentiate high- and low-affinity populations of postsynatpic .gamma.-aminobutyric acid (GABAA) receptors. The super-low-affinity GABAA receptors were characterizedby the enhancing effect of GABA on [3H]diazepam binding. The GABA antagonists 2-(3-carboxypropyl)-3-amino-4-methyl-6-phenylpyridazinium chloride (SR 95103) and 3-.alpha.-hydroxy-16-imino-5.beta.-17-aza-androstan-11-one (R 5135) shifted and suppressed the dose-response curve of GABA on diazepam binding. SR 95103 displaced the lower affinity [3H]GABA binding with higher potency. Dissociation of the binding of the antagonist 2-(3-carboxypropyl)-3-amino-6-p-methoxyphenylpyridazinium bromide ([3H]SR 95531) was polyphasic. Displacing potencies cfSR 95531 and GABAwere examined on the major (85%) rapid and minor slower phasses of dissociation separated kinetically. The slower phase corresponded to higher affinity binding of SR 95531 which was displaced by GABA with about 10 times less potency. Photoaffinity labeling with muscimol decreased the numbr of [3H]muscimol binding sites by 27%, but not that of bicuculline methiodide. These findings can be explained by a preferential binding of antagonists to hydrophobic accessory sites around low-afinity GABAA receptors.